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The experimental autoimmune encephalomyelitis (EAE) is used to study multiple sclerosis (MS) pathology and develop novel technologies to quantify inflammation over time. Magnetic resonance imaging (MRI) with gadolinium-based contrast agents (GBCAs) is the state-of-the-art method to assess inflammation in MS patients and its animal model. Fluorine (19F)-MRI is one novel technology to quantify inflammatory immune cells in vivo using 19F-nanoparticles. T1 mapping of contrast-enhancing images is another method that could be implemented to quantify inflammatory lesions. Transient macroscopic changes in the EAE brain confound quantification and necessitate registration methods to spatially align images in longitudinal studies. For 19F-MRI, an additional challenge is the low signal-to-noise ratio (SNR) due to low number of 19F-labeled immune cells in vivo. Transceive surface radiofrequency (RF) probes and SNR-efficient imaging techniques such as RARE (Rapid Acquisition with Relaxation Enhancement) are combined to increase sensitivity in 19F-MRI. However, the strong spatially-varying RF field (B1 inhomogeneity) of transceive surface RF probes further hampers quantification. Retrospective B1 correction methods typically use signal intensity equations, unavailable for complex acquisition methods like RARE. The main goal of this work is to investigate novel B1 correction and registration methods to enable the study of inflammatory diseases using 1H- and 19F-MRI following GBCA and 19F-nanoparticle administration, respectively. For correcting B1 inhomogeneities in 1H- and 19F-MR transceive surface RF probes, a model-based method was developed using empirical measurements and simulations, and then validated and compared with a sensitivity method and a hybrid of both. For 19F-MRI, a workflow to measure anatomical images in vivo and a method to compute 19F-concentration uncertainty after correction using Monte Carlo simulations were developed. To overcome the challenges of EAE brain macroscopic changes, a pipeline for registering images throughout longitudinal studies was developed. The proposed B1 correction methods demonstrated dramatic improvements in signal quantification and T1 contrast on images of test phantoms and mouse brains, allowing quantitative measurement with transceive surface RF probes. For low-SNR scenarios, the model-based method yielded reliable 19F-quantifications when compared to volume resonators. Uncertainty after correction depended linearly on the SNR (≤10% with SNR≥10.1, ≤25% when SNR≥4.25). The implemented registration approach provided successful image alignment despite substantial morphological changes in the EAE brain over time. Consequently, T1 mapping was shown to objectively quantify gadolinium lesion burden as a measure of inflammatory activity in EAE. The 1H- and 19F-MRI methods proposed here are highly relevant for quantitative MR of neuroinflammatory diseases, enabling future (pre)clinical investigations., Die experimentelle Autoimmun-Enzephalomyelitis (EAE) wird zur Untersuchung Multipler Sklerose (MS) und zur Entwicklung neuer Technologien zur Entzündungsquantifizierung eingesetzt. Magnetresonanztomographie (MRT) mit Gadolinium-haltigen Kontrastmitteln (GBCAs) ist die modernste Methode zur Beurteilung von Entzündungen bei MS-Patienten und im Tiermodell. Fluor (19F)-MRT unter Verwendung von 19F-Nanopartikeln ist eine neue Technologie zur Quantifizierung entzündlicher Immunzellen in vivo. T1-Kartierung ist eine MRT-Methode, die zur Quantifizierung entzündlicher Läsionen eingesetzt werden könnte. Temporäremorphologische Veränderungen im EAE-Gehirn erschweren die Quantifizierung und erfordern Registrierungsmethoden, um MRT-Bilder in Längsschnittstudien räumlichabzugleichen. Das niedrige Signal-Rausch-Verhältnis (SNR) ist aufgrund der geringen Anzahl 19F-markierter Immunzellen in vivo eine zusätzliche Herausforderung der 19F-MRT. Um deren Empfindlichkeit zu erhöhen, werden Sende-/Empfangsoberflächen-Hochfrequenzspulen (TX/RX-HF-Spule) und SNR-effiziente MRT-Techniken wie RARE (Rapid Acquisition with Relaxation Enhancement) kombiniert. Jedoch verhindert die starke räumliche Variation des HF-Feldes (B1-Inhomogenität) dieser Spulen die Signalquantifizierung. Retrospektive B1-Korrekturmethoden verwenden in der Regel Signalintensitätsgleichungen, die für komplexe MRT-Techniken wie RARE nicht existieren. Das Hauptziel dieser Arbeit ist die Untersuchung neuartiger B1-Korrektur- und Bildregistrierungsmethoden, um in vivo 1H- und 19F-MRT Studien von Entzündungsprozessen zu ermöglichen. Zur Korrektur von B1-Inhomogenitäten wurde eine modellbasierte Methode entwickelt. Diese verwendet empirische Messungen und Simulationen, wurde in Phantomexperimenten validiert und mit Referenzmethoden verglichen. Für 19F-MRT wurden ein Protokoll zur Messung anatomischer Bilder in vivo und eine Methode zur Berechnung der 19F-Konzentrationsunsicherheit nach Korrektur mittels Monte-Carlo-Simulationen entwickelt. Um morphologische Veränderungen im EAE-Gehirn in longitudinalen Studien zu kompensieren, wurde zur Bildregistrierung eine Software-Bibliothek entwickelt. Die B1-Korrekturmethoden zeigten in Testobjekten und Mäusehirnen drastische Verbesserungen der Signal- und T1 Quantifizierung und ermöglichten so quantitative Messungen mit TX/RX-HF-Spulen. Die modellbasierte Methode lieferte für geringe SNRs zuverlässige 19F-Quantifizierungen, deren Genauigkeit mit dem SNR korrelierte. Die implementierte Registrierungsmethode ermöglichte einen erfolgreichen Abgleich von Bildserientrotz erheblicher morphologischer Veränderungen im EAE-Hirn. Folglich wurde gezeigt, dass MRT basierte T1-Kartierung die Gadolinium-Läsionslast als Maß entzündlicher Aktivität bei EAE objektiv quantifizieren kann. Die hier unterscuhten Methoden sind für quantitative 1H- und 19F-MRT neuroinflammatorischer Erkrankungen sehr relevant und ermöglichen künftige (prä)klinische Untersuchungen.

Paramagnetic relaxation enhancement (PRE) is the current strategy of choice for enhancing magnetic resonance imaging (MRI) contrast and for accelerating MRI acquisition schemes. Yet, debates regarding lanthanides’ biocompatibility and PRE-effect on MRI signal quantification have raised the need for alternative strategies for relaxation enhancement. Herein, we show an approach for shortening the spin–lattice relaxation time (T1) of fluoride-based nanocrystals (NCs) that are used for in vivo 19F-MRI, by inducing crystal defects in their solid-crystal core. By utilizing a phosphate-based rather than a carboxylate-based capping ligand for the synthesis of CaF2 NCs, we were able to induce grain boundary defects in the NC lattice. The obtained defects led to a 10-fold shorter T1 of the NCs’ fluorides. Such paramagnetic-free relaxation enhancement of CaF2 NCs, gained without affecting either their size or their colloidal characteristics, improved 4-fold the obtained 19F-MRI signal-to-noise ratio, allowing their use, in vivo, with enhanced hotspot MRI sensitivity.

Purpose To assess alveolar perfusion by applying dynamic susceptibility contrast MRI to 19 F-MRI of inhaled perfluoropropane (PFP). We hypothesized that passage of gadolinium-based contrast agent (GBCA) through the pulmonary microvasculature would reduce magnetic susceptibility differences between water and gas components of the lung, elevating the T 2 ∗ of PFP. Methods Lung-representative phantoms were constructed of aqueous PFP-filled foams to characterize the impact of aqueous/gas phase magnetic susceptibility differences on PFP T 2 ∗ . Aqueous phase magnetic susceptibility was modulated by addition of different concentrations of GBCA. In vivo studies were performed to measure the impact of intravenously administered GBCA on the T 2 ∗ of inhaled PFP in mice (7.0 Tesla) and in healthy volunteers (3.0 Tesla). Results Perfluoropropane T 2 ∗ was sensitive to modulation of magnetic susceptibility difference between gas and water components of the lung, both in phantom models and in vivo. Negation of aqueous/gas phase magnetic susceptibility difference was achieved in lung-representative phantoms and in mice, resulting in a ~2 to 3× elevation in PFP T 2 ∗ (3.7 to 8.5 ms and 0.7 to 2.6 ms, respectively). Human studies demonstrated a transient elevation of inhaled PFP T 2 ∗ (1.50 to 1.64 ms) during passage of GBCA bolus through the lung circulation, demonstrating sensitivity to lung perfusion. Conclusion We demonstrate indirect detection of a GBCA in the pulmonary microvasculature via changes to the T 2 ∗ of gas phase PFP within directly adjacent alveoli. This approach holds potential for assessing alveolar perfusion by dynamic susceptibility contrast 19 F-MRI of inhaled PFP, with concurrent assessment of lung ventilation properties, relevant to lung physiology and disease.

Purpose Inflammation is involved in many disease processes. However, accurate imaging tools permitting diagnosis and characterization of inflammation are still missing. As inflamed tissues exhibit a high rate of glycolysis, pyruvate metabolism may offer a unique approach to follow the inflammatory response and disease progression. Therefore, the aim of the study was to follow metabolic changes and recruitment of inflammatory cells after onset of inflammation in arthritic ankles using hyperpolarized 1-13C-pyruvate magnetic resonance spectroscopy (MRS) and 19F magnetic resonance imaging (MRI), respectively. Procedure Experimental rheumatoid arthritis (RA) was induced by intraperitoneal injection of glucose-6-phosphate-isomerase-specific antibodies (GPI) containing serum. To monitor pyruvate metabolism, the transformation of hyperpolarized 1-13C-pyruvate into hyperpolarized 1-13C-lactate was followed using MRS. To track phagocytic immune cell homing, we intravenously injected a perfluorocarbon emulsion 48 h before imaging. The animals were scanned at days 1, 3, or 6 after GPI-serum injection to examine the different stages of arthritic inflammation. Finally, to confirm the pyruvate metabolic activity and the link to inflammatory cell recruitment, we conducted hematoxylin-eosin histopathology and monocarboxylase transporter (MCT-1) immune histochemistry (IHC) of inflamed ankles. Results Hyperpolarized 1-13C-pyruvate MRS revealed a high rate of lactate production immediately at day 1 after GPI-serum transfer, which remained elevated during the progression of the disease, while 19F-MRI exhibited a gradual recruitment of phagocytic immune cells in arthritic ankles, which correlated well with the course of ankle swelling. Histopathology and IHC revealed that MCT-1 was expressed in regions with inflammatory cell recruitment, confirming the metabolic shift identified in arthritic ankles. Conclusions Our study demonstrated the presence of a very early metabolic shift in arthritic joints independent of phagocytic immune cell recruitment. Thus, hyperpolarized 1-13C-pyruvate represents a promising tracer to monitor acute arthritic joint inflammation, even with minor ankle swelling. Furthermore, translated to the clinics, these methods add a detailed characterization of disease status and could substantially support patient stratification and therapy monitoring.

„Das Nicht-Wahrnehmen von etwas beweist nicht dessen Nicht-Existenz.“ Dalai Lama Im Gegensatz zum obigen Zitat beweist das Wahrnehmen von etwas dessen Existenz. Dieses Prinzip ist ein von Wissenschaftlern häufig angewendetes Prinzip und weist auf die Wichtigkeit von bildgebenden Verfahren für Medizin und life science hin. Bildgebende Verfahren, wie z.B. Fluorescence Imaging oder Magnetic Resonance Imaging, ermöglichen es Objekte oder Wechselwirkungen zwischen Objekten zu visualisieren, die im Interesse des Betrachters liegen. Allerdings kann eine bildgebende Technik nicht alle Daten eines Objektes vollständig liefern. Aufgrund dessen besteht die Möglichkeit zwei oder mehrere bildgebenden Techniken synergistisch zu kombinieren, um die Limitierung der einzelnen Methoden zu überschreiten. Im Fokus dieser Arbeit steht die Entwicklung von Molekülen, die gleichzeitig sowohl für das Fluorescence Imaging als auch für das Magnetic Resonanz Imaging geeignet sind. Als Fluoreszenzfarbstoffsystem dient hierbei die Farbstoffklasse der Boron-dipyrromethene (BODIPYs). Verschiedenste mono- bzw. difluorierte und trifluormethylierte BODIPY-Farbstoff wurden synthetisiert, um mit den erhaltenen Molekülen das erst seit einigen Jahrzehnten bekannte 19F-Magnetic Resonance Imaging durchzuführen. Zusätzlich wurde die Fragmentierung dieser BODIPYs mittels Massenspektroskopie untersucht. Die synthetisierten dualen Reporter zeigten gute Fluoreszenzeigenschaften, jedoch nur geringe 19F-Magnet-Resonanzaktivität. Aufgrund dessen wurden BODIPY-Farbstoffe mit 18 bzw. 27 magnetisch äquivalenten 19F-Atomen hergestellt. Diese hochfluorierten BODIPYs erwiesen sich als vielversprechende Kandidaten für das duale Fluorescence/ Magnetic Resonanz Imaging. “The non-perception of something does not prove its non-existence.” Dalai Lama Contrary to the quotation, the perception of something means its existence. This principle is often used by scientists and shows the importance of imaging modalities in medicine and life science. Imaging modalities, e.g. Fluorescence Imaging or Magnetic Resonance Imaging, allow visualizing objects of interest or interactions between different objects. However, a single imaging technique cannot provide all data of an object of interest. Hence, there exists the possibility to combine two or more imaging techniques to overcome the limitations of each single method. The focus of this work is the development of molecules which are suited for Fluorescence Imaging as well as Magnetic Resonance Imaging. The used chromophore scaffold for this project is the Boron-dipyrromethene (BODIPY). Several different mono- and difluororinated and trifluororinated BODIPY-dyes were synthesized to perform 19F-Magnetic Resonance Imaging, which is known for only a few decades. Additionally, the fragmentation of these BODIPYs was analyzed with mass spectroscopy. The synthesized dual reporters showed good fluorescence properties, but lacked 19F-magnetic resonance activity. As a result, BODIPY-dyes with 18 respectively 27 magnetic equivalent 19F-atoms were synthesized. These high fluorinated BODIPYs turned out to be promising candidates for dual fluorescence/ magnetic resonance imaging.

Neural stem cell implantations have been extensively investigated for treatment of brain diseases such as stroke. In order to follow the localization and functional status of cells after implantation noninvasive imaging is essential. Therefore, we developed a comprehensive multi-modality platform for in vivo imaging of graft localization, density, and survival using 19F magnetic resonance imaging in combination with bioluminescence imaging. We quantitatively analyzed cell graft survival over the first 4 weeks after transplantation in both healthy and stroke-damaged mouse brain and correlated our findings of graft vitality with the host innate immune response. The multi-modality imaging platform will help to improve cell therapy also in context other than stroke and to gain indispensable information for clinical translation.

Graphical abstract A water-soluble 1,1-geminal fluorinated bisphosphonate showing a single and narrow 19F resonance was synthesised and characterised. Its properties as contrast agent for 19F-MRI were evaluated in vitro and in vivo., Highlights • A trifluoromethyl-bisphosphonate for 19F-MRI was synthesised and characterised. • In vitro studies showed its potential with properties comparable to previous probes. • In vivo 19F/1H-MRI studies showed fast renal excretion and liver uptake., 19F-magnetic resonance imaging (MRI) is a promising technique that may allow us to measure the concentration of exogenous fluorinated imaging probes quantitatively in vivo. Here, we describe the synthesis and characterisation of a novel geminal bisphosphonate (19F-BP) that contains chemically-equivalent fluorine atoms that show a single and narrow 19F resonance and a bisphosphonate group that may be used for labelling inorganic materials based in calcium phosphates and metal oxides. The potential of 19F-BP to provide contrast was analysed in vitro and in vivo using 19F-MRI. In vitro studies demonstrated the potential of 19F-BP as an MRI contrast agent in the millimolar concentration range with signal-to-noise ratios (SNR) comparable to previously reported fluorinated probes. The preliminary in vivo MRI study reported here allowed us to visualise the biodistribution of 19F-BP, showing uptake in the liver and in the bladder/urinary system areas. However, bone uptake was not observed. In addition, 19F-BP showed undesirable toxicity effects in mice that prevent further studies with this compound at the required concentrations for MRI contrast. This study highlights the importance of developing 19F MRI probes with the highest signal intensity achievable.

19F-magnetic resonance imaging (MRI) is a promising technique that may allow us to measure the concentration of exogenous fluorinated imaging probes quantitatively in vivo. Here, we describe the synthesis and characterisation of a novel geminal bisphosphonate (19F-BP) that contains chemically-equivalent fluorine atoms that show a single and narrow 19F resonance and a bisphosphonate group that may be used for labelling inorganic materials based in calcium phosphates and metal oxides. The potential of 19F-BP to provide contrast was analysed in vitro and in vivo using 19F-MRI. In vitro studies demonstrated the potential of 19F-BP as an MRI contrast agent in the millimolar concentration range with signal-to-noise ratios (SNR) comparable to previously reported fluorinated probes. The preliminary in vivo MRI study reported here allowed us to visualise the biodistribution of 19F-BP, showing uptake in the liver and in the bladder/urinary system areas. However, bone uptake was not observed. In addition, 19F-BP showed undesirable toxicity effects in mice that prevent further studies with this compound at the required concentrations for MRI contrast. This study highlights the importance of developing 19F MRI probes with the highest signal intensity and lowest toxicity achievable.

A protocol to load perfluoro-crown-ether (PFCE) nanoemulsion directly into yeast-derived glucan particles (GPs) was developed. It was observed that the PFCE encapsulation did not affect the 19F-MRI properties of the nanoemulsion that is currently in clinical trials. GPs loaded with PFCE nanoemulsion were taken up avidly by murine macrophages in vitro, resulting in a cellular uptake 150 % higher than the not GPs-entrapped nanoemulsion. Accordingly, a corresponding improvement in the 19F-MRI detection of the labelled cells can be obtained. The high biotolerability and versatility of GPs, make these microcarriers a promising option for designing of improved in vivo cellular imaging protocols.

Perfluorocarbon (PFC) particles are currently on the rise as cell labeling agents for ¹⁹F-MRI tracking of dendritic cell (DC)-based vaccines. In this work, we design theranostic PFC particles for single-step loading of DCs with both antigenic protein and with a liquid PFC for ¹⁹F-MRI detection of the antigen-loaded cells. Upon addition to DCs in vitro, the antigen-loaded PFC particles are efficiently internalized, resulting in intracellular presence of up to 40 pmol ¹⁹F atoms per cell. At the same time, the DCs become loaded with antigenic proteins, that can be efficiently processed, without important effects on cell viability or altering the DC's phenotype and the cell's capacity to respond to danger signals. In addition, antigen-loaded PFC particle containing DCs are capable of inducing extensive proliferation of antigen-specific CD8⁺ T cells in vitro. Importantly, the antigen-coated PFC particles allow in vitro ¹⁹F-MRI-based detection of the antigen-containing DCs with detection limits as low as 10³ cells μl⁻¹. The dual-modality characteristics of the designed particles could assure that only those DCs that have taken up the antigen, and hence are responsible for an immune response, are traceable via ¹⁹F-MRI. Taken together, these novel dual-modality particles represent an interesting strategy in the development of a traceable DC vaccine.

Cyclam (1,4,8,11-tetraazacyclotetradecane) derivatives are widely used for various purposes. In a medicine they are applied as ligands for radiometals applied as diagnostic or therapeutic agents against tumours, hypoxic brain tissues etc. In this thesis a new method for asymmetric cyclam derivati- ves preparation was developed. New cyclam derivatives with trifluoroethyl groups were prepared too. Paramagnetic metal complexes with these fluori- nated ligands can be used as contrast agents for 19 F-MRI. Keywords: Cyclam; Non-symmetric protection; Contrast agents; Trifluoroethylamines; Paramagnetic relaxation; 19 F-MRI.